Polysaccharide sulfates
I. Conversion of sodium sulfates of starch, carboxymethylstarch,
and (4-0-methyl-D-glucurono)-D-xylan into hydrogen sulfates
K. TIHLÁRIK and E. LATTOVÁ
institute of Chemistry, Slovak Academy of Sciences, CS-84238 Bratislava
Received 2 August 1989
Sodium sulfates of starch, carboxymethylstarch, and (4-0-methyl-D-glucurono)-D-xylan were prepared using the sulfur trioxide—pyridine complex or chlorosulfonic acid. Conversion of derivatives prepared into the
corresponding hydrogen sulfates was carried out with hydrochloric acid in
isopropyl alcohol in the presence of a little amount of water. Approximately
20 and 3 % decrease in the sulfur content was observed with derivatives of
starch and (4-0-methyl-D-glucurono)-D-xylan, respectively, on conversion
under the given conditions; the latter revealed a substantially lower elimination of the sulfate group. Determination of acid groups by differential
Potentiometrie method (titration with sodium methoxide in dimethyl sulfoxide) afforded a little lower values than those of methods employed so far.
Polysaccharide sulfates are a noticeable group of derivatives utilizable in
various branches; derivatives of starch and (4-0-methyl-D-glucurono)-D-xylan
are valuable especially in pharmacology. Investigated were physiological effects
of starch sulfates and their components aiming to prepare a substitute for the
natural anticoagulant heparin [1—3]. (4-0-Methyl-D-glucurono)-D-xylan sulfates were prepared to examine the relationship between their structure and
antithrombotic or antilipemic activities [4].
Sulfatation of the above-mentioned polysaccharides with sulfuric acid is
unfavourable because of considerable degradation of the substrate especially of
starch [5]. More advantageous is the treatment with chlorosulfonic acid in a
suitable solvent, as e.g. in pyridine [6], or in the presence of 7V,7V-dimethylformamide [7]. Polysaccharides can also be sulfatized with complexes of sulfur
trioxide in a suited solvent, as e.g. pyridine, triethylamine, 7V,7V-dimethylformamide or dioxane [8].
Polysaccharide sulfates are known to be stable in the form of their sodium
salts. Their conversion into hydrogen sulfates is associated with some problems.
Thus, Whistler and Spencer [5] reported that some hydrogen sulfates of polysaccharides undergo an easy autohydrolysis due to a high dissociation of the
functional group. According to Japanese authors [9] the dissociation degree of
the hydrogen sulfate group of cellulose hydrogen sulfate derivative equals 1
Chem. Papers 45 (4) 547—552 (1991)
547
K. T1HLÁRIK, E. LATT0VÁ
irrespectively of the pH of the medium in the actual region; on the other hand,
the dissociation degree of the carboxyl group in the afore-mentioned type of
derivatives is pH dependent.
This paper concerns the examination of sodium polysaccharide sulfates
conversion (of starch, carboxymethylstarch, and (4-0-methyl-D-glucurono)-D-xylan) to hydrogen sulfates. The possibility to determine the acid groups by
differential Potentiometrie titration in a nonaqueous medium was verified.
Experimental
Potato starch and carboxymethylstarch (vv(COCH3) = 8.65 %) are technical products
(České škrobárny, Havlíčkův Brod), (4-0-methyl-D-glucurono)-D-xylan (Mn = 18 000,
u'(4-0-methyl-D-glucuronic acid) = 17.30 %) was isolated from beech wood according to
[10]. The low-molecular xylan (Mn = 8000, vv(4-0-methyl-D-glucuronic acid) = 17.20 %)
was prepared from the above-mentioned product according to [11]. Solution of 0.1 M
sodium methoxide was obtained according to [12] in the mixture methanol—dimethyl
sulfoxide (<pr = 1:20). 4 M-HC1 was prepared by mixing concentrated hydrochloric acid
(37%) and isopropyl alcohol in the appropriate ratio. The sulfur trioxide—pyridine
complex is a commercial product (Merck, FRG).
Determination of sulfur after mineralization of samples was done gravimetrically
through barium sulfate according to [13]. The content of uronic acids was determined by
the carbazole method according to [14]. All samples were dried under reduced pressure
prior to determination at 60 °C for 5 h. Digital pH-meter OP-208/1 (Radelkis, Budapest)
equipped with antimony electrode of a usual type and a calomel electrode in which the
saturated aqueous KCl solution was replaced by a saturated methanolic KCl solution
was employed for Potentiometrie titrations.
Starch and carboxymethylstarch
sulfates
3
Suspension of the respective polysaccharide (4g) in A^,7V-dimethylformamide (50 cm )
was added to the sulfur trioxide—pyridine complex (8g) in 7V,N-dimethylformamide
3
(100 cm ) at room temperature at which the mixture was shaken for 16h. Aqueous
3
solution of sodium hydrogencarbonate (50 cm ; 10 %) was added to the product; metha­
3
nol (100 cm ) was introduced into the stirred suspension, the product was filtered off and
dried. Yield = 4.10 g and 4.05 g of starch and carboxymethylstarch sulfates, respectively.
(4-0-Methyl-D-glucurono)'D-xylan
sulfates
To (4-Omethyl-D-glucurono)-D-xylan (6g) dissolved in N,7V-dimethylformamide
3
3
3
(200 cm ) a mixture of pyridine (32.5 cm ) and chlorosulfonic acid (8.1 cm ) was dropwise
548
Chem. Papers 45 (4) 547—552 (1991)
POLYSACCHARIDE SULFATES. I
added with stirring at 0°C. The mixture was then heated to 75 °C and stirred for 4.5 h,
cooled and poured into ethanol (2.2 dm3). The precipitate was filtered off, washed three
times with ethanol (100 cm3 each) and dissolved in distilled water (500 cm3). The pH of
the solution was adjusted with 1 M-NaOH to 8.8, the solution was concentrated to ca.
120cm3 and the product was precipitated with acetone—ethanol (<pr = 3: 1; 2.5dm3).
The lightbrown precipitate was filtered off, dissolved in distilled water (100 cm3) and the
solution was left to dialýze against distilled water for 72 h. Yield = 6.5 g, A?n = 11 500.
For sulfatation of the low-molecular xylan the same procedure was applied. The yield
from 0.60 g of the original material was 0.62 g, Mn = 4900.
Conversion of sodium sulfates of polysaccharides to the corresponding hydrogen
sulfates
A solution of 4M hydrochloric acid in isopropyl alcohol (10cm3) was added to a
stirred suspension of sodium polysaccharide sulfate (1 g) in isopropyl alcohol (50 cm3).
After 5 min stirring in a stoppered flask the suspension was filtered off and washed with
methanol till the absence of chloride ions. The product was dried and the content of
hydrogen sulfate and carboxyl groups was titrimetric.ally determined immediately.
Determination of acid groups in hydrogen sulfates by differential
titration
The sample (50 mg) was dissolved in dimethyl sulfoxide (20 cm3) in a stoppered flask.
The air was removed by washing with gaseous nitrogen for 10 min and the content was
determined by titration with 0.1 M sodium methoxide added in 0.1 cm3 portions by
means of Potentiometrie curve reading.
Results and discussion
Starch and carboxymethylstarch were sulfatized by treatment with the sulfur
trioxide—pyridine complex. The required substitution degree corresponding to
4.0—6.5 mass % of sulfur was achieved in 7V,A^-dimethylformamide during 16 h
at room temperature. The substitution degree was lower when applying pyridine
as a solvent.
(4-0-Methyl-D-glucurono)-D-xylan prepared according to [10] was depolymerized according to [11], because relative molecular mass of the product
required for sulfatation should not exceed 10000. The so-called low-molecular
xylan thus obtained was reacted under the same conditions.
Conversion of sodium polysaccharide sulfates to the corresponding hydrogen
sulfates is accompanied with autohydrolysis [5]. Also the hydrogen sulfate
Chem. Papers 45 (4) 547—552 (1991)
549
Table 1
Determination of sulfur and uronic acid in sulfated polysaccharides
vi'(uronic acid)/%
w(S)/%
Parent polysaccharide
Potato starch
Carboxymethylstarch
xylan
(4-0-Methyl-D-glucurono)-D-xylan
Low-molecular xylan
Sulfate
Hydrogen sulfate form
Sulfate
Hydrogen sulfate form
via BaS04
via BaS04
MeONa
Carbazole
Carbazole
MeONa
6.50
4.30
14.20
16.10
4.82
3.22
13.80
15.60
4.75
3.03
13.28
15.10
—
—
—
—
—
15.60
10.50
15.10
10.05
14.50
9.90
POLYSACCHARIDE SULFATES. I
group is very reactive and therefore we investigated several conversion
procedures. The most favourable agent was found to be hydrochloric acid in
isopropyl alcohol.
The products were characterized by routine methods employed for sulfur and
uronic acid determinations. At the same time we tried to differentiate and
determine the acid groups by Potentiometrie titration with sodium methoxide in
yV,7V-dimethylformamide. The results are presented in Table 1.
It has been found that conversion of sodium polysaccharide sulfates (values
for the starting sulfates in Table 1 are reduced to H + form to be comparable)
into hydrogen sulfates was accompanied by a decrease in the sulfur content in
starch and carboxymethylstarch by ca. 20%, which means a considerable elimination of the functional group; on the other hand, with (4-O-methyl-D-glucurono)-D-xylan derivatives only a 3 % decrease of the sulfur content occurred.
Some analogy might be seen with the alkaline desulfatation of some polysaccharide derivatives, where elimination of sulfate groups proceeds more easily
when the saccharide unit of the polysaccharide bore a free hydroxyl group at
C-6 [5]; this is, however, not the case with xylan derivatives.
Routine method was employed for titrimetric determination of hydrogen
sulfate samples with sodium methoxide in dirhethyl sulfoxide. (Evaluation of
Potentiometrie curves will be presented separately.) With starch derivatives the
hydrogen sulfate group was estimated for 10 parallel titrations. The calculated
standard deviation s and variation coefficient were 0.326 and 7.43 %, respectively. Comparison of results obtained by gravimetric and titration methods of
sulfur determination showed a little lower values with the second one.
Content of uronic acid was examined in both xylan types of compounds; as
found no considerable decarboxylation occurred under the given conditions.
Values obtained by titration are again but unsignificantly lower than those of
carbazole method.
References
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Translated by Z. Votický
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